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Bovine and Porcine Adipogenesis, Myogenesis, and Tissue Engineering Strategies to Improve Flavor and Pigmentation of Cell-Based Meat

Krieger, Jessica
http://rave.ohiolink.edu/etdc/view?acc_num=kent160657063728198

Abstract Details

2020, PHD, Kent State University, College of Arts and Sciences / School of Biomedical Sciences.
Background: Animal agriculture is an industrialized, globalized system of meat production that will continue to increase in demand through the 21st century. This is a greatly resource intensive process that produces greenhouse gases and zoonotic disease development that contribute to climate change and public health pandemics. Since the demand for meat shows no sign of slowing in the coming years, alternative methods of meat production are required to feed a growing human population. Cell-based meat is one possible solution, which is meat grown from cell culture technology. Cell-based meat can be grown from developing edible cell lines and expanding them in bioreactors using biomedical techniques and equipment. The field in in its infancy, however, and many questions remain about how to develop useful cell lines and which are the most effective ways to grow them. This work demonstrates a research framework from which to characterize and compare two of the main cell types in meat: skeletal muscle cells and intramuscular fat cells. Methods were developed to analyze the difference between porcine and bovine cells, what characterizes their sensory and pigment properties, and how to efficiently grow them skeletal muscle tissue form. Methods: Bovine and porcine myoblasts and intramuscular fibroblast were isolated from the hind leg of 2 month old pigs and cows. Myoblasts were used for myogenesis assays, and intramuscular fibroblasts were used for adipogenesis assays. Gene and protein expression and volumetric hypertrophy data was obtained for myoblasts and lipid staining was quantified for fibroblasts. Meat color and pigment was determined for pork, beef, and cultured cells using colorimetry. Myoglobin gene and protein expression was assessed during myogenesis assays with and without electrical stimulation, mimicking an exercise regimen. Conditioned media from polarized porcine macrophages was used to characterize the potential for developing serum free media for myoblast proliferation and differentiation. Self-assembly, hydrogel-based, and plant-scaffold based methods of tissue engineering were performed with C2C12s and porcine myoblasts to identify methods useful for scaling up the production of meat tissue. Results: We found that that while bovine and porcine myoblasts undergo similar patterns of myogenic transcription factor gene expression, they undergo different degrees of hypertrophy, with bovine cells increasing in volume by ~40% more than porcine myoblasts. Bovine adipocytes were also less mature than porcine adipocytes but were higher in population frequency. C2C12 cells were found to be white in color compared to the deep red found in oxygenated beef and pinkish red found in pork. Myoglobin from beef samples was higher than in culture bovine myocytes, and myoglobin could be increased through electrical stimulation regimen of bovine myocytes. Macrophage conditioned media improved proliferation and differentiation in M0 and M2 phenotypes, indicating that a media producing cell line could be useful for producing proliferation and differentiation media for myogenic cultures. C2C12s were easily incorporated into self-assembled and hydrogel-based tissues, but porcine myoblast tissues were unable to keep integrity over long term culture. C2C12s were also seeded into cellulose-based scaffolds derived from celery. Cells took several weeks to adhere to cellulose scaffolds and differentiate into myotubes. Subsequently, porcine fibroblast and myofibroblast feeder layer studies indicated that myofibroblast feeder layers may be a useful option for prepping cellulose scaffolds for myoblast adhesion. Discussion: Bovine and porcine cells may yield different biomasses in bioprocesses designed to produce cell based meat. They may also present different myoglobin protein expression levels in vitro, which would require different research strategies to amplify protein concentration. Electrical stimulation regimens are one strategy to increase myoglobin protein in a bioprocess, which would enhance flavor and pigment of cells. Additionally, an immortal cell line could be developed that could produce a growth factor cocktail that would be suitable for supplying serum free media to an edible cell line. Tissue engineering whole cuts of meat is many decades away, but the techniques outlined in this work could be useful for developing microcarrier platforms for scale up of cell-based meat.
Min-Ho Kim, Dr. (Advisor)
Kristy Welshhans, Dr. (Committee Member)
Oleg Lavrentovich, Dr. (Committee Member)
Feng Dong, Dr. (Committee Member)
Songping Huang, Dr. (Committee Member)
121 p.
Biomedical Engineering; Biomedical Research; Food Science
Food science; meat science; bioengineering; biomedical engineering; biomedical sciences; in vitro meat; cell-based meat; cultured meat; cultivated meat; myogenesis; adipogenesis; bovine; porcine; myoglobin

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Citations

  • Krieger, J. (2020). Bovine and Porcine Adipogenesis, Myogenesis, and Tissue Engineering Strategies to Improve Flavor and Pigmentation of Cell-Based Meat [Doctoral dissertation, Kent State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=kent160657063728198

    APA Style (7th edition)

  • Krieger, Jessica. Bovine and Porcine Adipogenesis, Myogenesis, and Tissue Engineering Strategies to Improve Flavor and Pigmentation of Cell-Based Meat . 2020. Kent State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=kent160657063728198.

    MLA Style (8th edition)

  • Krieger, Jessica. "Bovine and Porcine Adipogenesis, Myogenesis, and Tissue Engineering Strategies to Improve Flavor and Pigmentation of Cell-Based Meat ." Doctoral dissertation, Kent State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=kent160657063728198

    Chicago Manual of Style (17th edition)

kent160657063728198
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This open access ETD is published by Kent State University and OhioLINK.